Our overall goal is to understand how genetic alterations in synovial sarcoma affect its cellular phenotype and clinical behavior. We have found that the type of fusion transcript (SYT-SSX1 vs. SYT-SSX2) resulting from the characteristic t (x;18) in synovial sarcomas is strongly correlated with tumor phenotype (monophasic vs, biphasic histology) and may also be associated with differences in clinical course. Our general hypothesis is that functional differences between SYT-SSX1 and SYT- SSX2, two chimeric transcriptional regulators presumed to function through protein-protein interactions, lead to differences in target gene expression and ultimately cellular phenotype, thereby defining distinct biological and clinical subgroups in synovial sarcoma. Secondary genetic alterations in G1/S control may determine another level of clinical subgroups in synovial sarcoma. Secondary genetic alterations in G1/S control may determine another level of clinical heterogeneity. AIM 1. Prognostic impact of SYT-SSX fusion type in synovial sarcoma. In series of 39 patients with localized synovial sarcoma. Secondary genetic alterations in G1/S control may determine another level of clinical heterogeneity. AIM 1. Prognostic impact of SYT-SSX fusion type in synovial sarcoma. In series of 39 patients w3ith localized synovial sarcoma, we found that cases with the SYT-SSX2 fusion that a significantly better metastasis free survival than cases with SYT-SSX1. To confirm this, we will collect data on an unrelated, larger, and more homogenous study group. A multicenter retrospective design will be used to address this question in a reasonable time frame. AIM 2. Prognostic impact of secondary genetic alterations in synovial sarcoma. Preliminary data from our collaborating group (Carlos Cordon-Cardo) suggest that P53 alteration is prognostically significant in synovial sarcoma. P53 mutation and other secondary genetic alterations potentially critical to G1/S control have so far not been systematically characterized in this tumor. We will complete the analysis of these genetic alterations prospectively in new cases and in our retrospective series. We hypothesize that P53 mutation and other genetic lesions affecting G1/S control will identify a small subset of patients with a markedly unfavorable prognosis, independent of SYT-SSX fusion type. AIM 3. Differential gene expression associated with alternative SYT-SSX fusion proteins, The association of SYT-SSX type and morphologic epithelial differentiation (biphasic histology) suggests that putative functional differences between ST-SSX1 and SYT-SSX2 may result in differences in the expression of direct or indirect target genes involved in this process. The transcripts to be analyzed for differential expression will come from three sources. 1. selected candidate genes involved in mesenchymal-epithelial transition; 2. putative differentially expressed transcripts isolated in our differential display studies; 3. putative differentially expressed transcripts identified collaboratively through large scale cDNA microarray analyses performed by Paul Meitzer's group at NHGRI on our primary tumor RNAs with either SYT-SSX1 or SYT- SSX2. Differential expression will be confirmed using several independent approaches. AIM 4. Analysis of SYT-SSX1 and SYT-SSX2 genomic breakpoints. The ratio of SSX1 to SSX2 breakpoints in synovial sarcoma is approximately 2:1. Our preliminary work shows that the introns are of approximately equal size, but may differ in the presence of repetitive elements. To further our understanding of the mechanistic basis of the t(x;18) translocation in synovial sarcoma, we will analyze the sequences of the involved SSX1 and SSX2 introns, and of the genomic translocation junctions in cases of synovial sarcomas with SYT-SSX1 or SYT-SSX2, respectively, for previously described putative junctions in cases of synovial sarcomas with SYT-SSX1 or SYT-SSX2, respectively, for previously described putative recombinogenic elements and clustering of breakpoints. We hypothesize that the involved SSX1 intron contains recombinogenic elements absent from the SSX2 intron, accounting for the uneven incidence of breakpoints.